Previous studies have demonstrated that exposure of mammalian cells to chromate resulted in the formation of DNA-protein crosslinks. A high percentage of protein complexed with DNA is mediated through linkage with Cr(III), however, some of the DNA-protein crosslinks are also mediated by oxidative components. This hypothesis is based upon the findings that during the reduction of Cr(VI) to Cr(III) oxygen radicals were formed. Previous studies have demonstrated that agents which generate oxygen radicals, such as x-rays, produce DNA-protein crosslinks. In the present grant, we will characterize the proteins that are complexed to the DNA through a non-chelatable linkage (i.e., oxidation) using intact Chinese hamster ovary cells. We will also determine the proteins that are complexed to DNA (as a background) in untreated cells. Attempts will be made through the use of antibodies and known standards to identify the proteins that are complexed to the DNA by each proposed linkage. Focus on identification of proteins will be based on those common to both or abundant proteins that may be unique to each process. We will determine whether chromate-induced DNA-protein crosslinks occur preferentially in transcriptionally active or inactive DNA since a variety of evidence suggests that it is likely to favor formation in transcriptionally active DNA. Finally, we will determine the DNA bases that are modified following treatment of intact cells with Cr(VI). Focus will be placed upon bases containing 51Cr, as well as those with amino acid linkages. These studies will aid in elucidating the molecular mechanisms by which chromate produces a major DNA lesion, the DNA- protein crosslink. DNA-protein crosslinks could represent an important lesion in the process of carcinogenesis, since they are not readily repaired and represent a bulky lesion that may have significant mutagenic consequences.